Phosphono-hydroisoquinoline compounds useful in reducing neurotoxic injury

ABSTRACT

A class of phosphono-hydroisoquinoline compounds is described for treatment to reduce neurotoxic injury associated with anoxia or ischemia which typically follows stroke, cardiac arrest or perinatal asphyxia. The treatment includes administration of a phosphono-hydroisoquinoline compound alone or in a composition in an amount effective as an antagonist to inhibit excitotoxic actions at major neuronal excitatory amino acid receptor sites. Compounds of most interest are those of the formula: ##STR1## wherein each of R 1  through R 4  is hydrido, each of Z 1  and Z 2  is hydroxyl and wherein the A ring is saturated.

This is a continuation of application Ser. No. 07/654,852 filed Feb. 13,1991, now abandoned, which is a division of 07/260,839 filed Oct. 21,1988 and issued as U.S. Pat. No. 4,997,821.

FIELD OF THE INVENTION

This invention is in the field of clinical neurology and relatesspecifically to a class of compounds, compositions and methods forneuro-protective purposes such as controlling chronic or acuteneurotoxic injury or brain damage resulting from neuro-degenerativediseases. For example, these compounds are particularly useful fortreating neurotoxic injury which follows periods of anoxia or ischemiaassociated with stroke, cardiac arrest or perinatal asphyxia. Thecompounds would also be useful as anti-convulsants and analgesics.

BACKGROUND OF THE INVENTION

Unlike other tissues which can survive extended periods of hypoxia,brain tissue is particularly sensitive to deprivation of oxygen orenergy. Permanent damage to neurons can occur during brief periods ofhypoxia, anoxia or ischemia. Neurotoxic injury is known to be caused oraccelerated by certain excitatory amino acids (EAA) found naturally inthe central nervous system (CNS). Glutamate (Glu) is an endogenous aminoacid which has been characterized as a fast excitatory transmitter inthe mammalian brain. Glutamate is also known as a powerful neurotoxincapable of killing CNS neurons under certain pathological conditionswhich accompany stroke and cardiac arrest. Normal glutamateconcentrations are maintained within brain tissue by energy-consumingtransport systems. Under low energy conditions which occur duringconditions of hypoglycemia, hypoxia or ischemia, cells can releaseglutamate. Under such low energy conditions the cell is not able to takeglutamate back into the cell. Initial glutamate release stimulatesfurther release of glutamate which results in an extracellular glutamateaccumulation and a cascade of neurotoxic injury.

It has been shown that the sensitivity of central neurons to hypoxia andischemia can be reduced by either blockage of synaptic transmission orby specific antagonism of postsynaptic glutamate receptors [see S. M.Rothman and J. W. Olney, "Glutamate and the Pathophysiology ofHypoxia--Ischemic Brain Damage", Annals of Neurology, Vol. 19, No. 2(1986)]. Glutamate is characterized as a broad spectrum agonist havingactivity at three neuronal excitatory amino acid receptor sites. Thesereceptor sites are named after the amino acids which selectively excitethem, namely: Kainate (KA), N-methyl-D-aspartate (NMDA or NMA) andquisqualate (QUIS). Glutamate is believed to be a mixed agonist capableof binding to and exciting all three receptor types.

Neurons which have EAA receptors on their dendritic or somal surfacesundergo acute excitotoxic degeneration when these receptors areexcessively activated by glutamate. Thus, agents which selectively blockor antagonize the action of glutamate at the EAA synaptic receptors ofcentral neurons can prevent neurotoxic injury associated with anoxia,hypoxia or ischemia caused by stroke, cardiac arrest or perinatalasphyxia.

Aminophosphonic acids have been investigated as neurotransmitterblockers [see M. N. Perkins et al, Neuroscience Lett., 23, 333 (1981);and J. Davies et al, Neuroscience Lett., 21, 77 (1981)]. In particular,compounds such as 2-amino-4-(2-phosphonomethylphenyl)butyric acid and2-(2-amino-2-carboxy)ethylphenylphosphonic acid have been synthesizedfor evaluation as antagonists in blocking the action of theneurotransmitter compounds L-glutamic acid and L-aspartic acid [K.Matoba et al, "Structural Modification of Bioactive Compounds II.Syntheses of Aminophosphonic Acids", Chem. Pharm. Bull., 32, (10)3918-3925 (1984)].

U.S. Pat. No. 4,657,899 to Rzeszotarski et al describes a class ofω-[2-(phosphonoalkylenyl)phenyl]-2-aminoalkanoic acids characterized asbeing selective excitatory amino acid neurotransmitter receptorblockers. These compounds are mentioned for use as anticonvulsants,antiepileptics, analgesics and cognition enhancers. Typical compounds ofthe class include 3-[2-phosphonomethylphenyl]-2-aminopropanoic acid and3-[2-(2-phosphonoethyl)phenyl]-2-aminopropanoic acid. European PatentApplication 203,891 of Hutchison et al. describes phosphonoalkylsubstituted pipecolic acid derivatives useful for treatment of nervoussystem disorders in mammals and as antagonists of the NMDA sensitiveexcitatory amino acid receptor, an example of which iscis-4-phosphonomethyl-2-piperidine carboxylic acid. West German PatentApplication 3,736,016 of Sandoz describes phosphonoalkyl phenylglycinesderivatives useful as anticonvulsant and as antagonists of the NMDAreceptor, an example of which is 3-(phosphonomethyl)phenylglycine. U.S.application Ser. No. 111,749 filed Oct. 21, 1987 describes certainphosphonoalkylphenylglycine derivatives useful in reducing neurotoxicinjury and as anticonvulsants and analgesics, an example of which is4-(phosphonomethyl)phenylglycine.

Other classes of compounds have been tested as agonists in blockingNMDA- or KA-induced neurotoxicity [J. W. Olney et al., "TheAnti-Excitotoxic Effects of Certain Anesthetics, Analgesics andSedative-Hypnotics", Neuroscience Letters, 68, 29-34 (1986)]. The testedcompounds included phencylidine, ketamine, cyclazocine, kynurenate andvarious barbiturates such as secobarbital, amobarbital andpentobarbital.

DESCRIPTION OF THE INVENTION

Control of neuropathological processes and the neurodegenerativeconsequences thereof in mammals is provided by treating a mammalsusceptible to neurologic injury with a compound of a classcharacterized in having activity as antagonists at a major neuronalexcitatory amino acid receptor site. This class of NMDA antagonistcompounds is also expected to contain compounds having anti-convulsantand analgesic activity. Such NMDA antagonist compounds may be selectedfrom a class of phosphono-hydroisoquinoline compounds defined by FormulaI: ##STR2## wherein each of R¹ through R³ is independently selected fromhydrido, alkyl, haloalkyl, halo, cyano, nitro and groups represented by--OR⁵, --SR⁵, ##STR3## wherein R⁵ is selected from hydrido, alkyl, aryland aralkyl; and wherein R⁴ is selected from hydrido, alkyl, acyl,aralkyl and ##STR4## and wherein each of Z¹ and Z² is independentlyselected from --OR⁵, SR⁵, ##STR5## wherein R⁵ is defined as before; andwherein the A ring can be either saturated, partially unsaturated orfully unsaturated, i.e., an aromatic ring. Within this class ofphosphono-hydroisoquinolines of the invention are the pharmaceuticallyacceptable salts of the compounds of Formula I, including acid additionsalts, base addition salts including alkali metal salts. Also includedwithin this class of compounds of the invention are tautomeric forms ofthe defined compounds and isomeric forms including diastereomers andenantiomers.

A preferred class of compounds within Formula I consists of thosecompounds wherein each of R¹ to R³ is independently selected fromhydrido, alkyl, halcalkyl, halo, cyano, nitro, --OR⁵ and --SR⁵ ; whereinR⁵ is selected from hydrido, alkyl, aryl and aralkyl; and wherein R⁴ isselected from hydrido, alkyl, acyl, aralkyl and --COOR⁵ ; wherein eachof Z¹ and Z² is independently selected from --OR⁵, --SR⁵, NR⁴ R⁵ and--OCHR⁵ OCOR⁵ ; and wherein the A ring can be either saturated,partially unsaturated or fully unsaturated (aromatic).

A more preferred class of compounds within Formula I consists of thosecompounds wherein each of R¹ to R³ is independently selected fromhydrido, alkyl, haloalkyl, halo, cyano, --OR⁵, wherein R⁵ is selectedfrom hydrido and alkyl; wherein R⁴ is selected from hydrido, alkyl,acyl, aralkyl and --COOR⁵ ; wherein each of Z¹ and Z² is independentlyselected from --OR⁵, NR⁴ R⁵ and --OCHR⁵ OCOR⁵ ; and wherein the A ringcan be either saturated, partially unsaturated or fully unsaturated(aromatic).

An even more preferred class of compounds within Formula I consists ofthose compounds wherein each of R¹, R² and R³ is hydrido; wherein R⁴ isselected from hydrido, alkyl, acyl, aralkyl and --COOR⁵ ; wherein R⁵ isselected from hydrido and alkyl; wherein each of Z¹ and Z² isindependently selected from --OR⁵, NR⁴ R⁵ and --OCHR⁵ OCOR⁵ ; andwherein the A ring can be either saturated, partially unsaturated orfully unsaturated (aromatic).

A more highly preferred class of compounds within Formula I consists ofthose compounds wherein each of R¹, R² and R³ is hydrido; wherein R⁴ isselected from hydrido, acyl and --COOR⁵ ; wherein R⁵ is selected fromhydrido and alkyl; wherein Z¹ is selected from --OR⁵, NR⁴ R⁵ and --OCHR⁵OCOR⁵ ; wherein Z² is hydroxyl; and wherein the A ring can be eithersaturated, partially unsaturated or fully unsaturated (aromatic).

A still more highly preferred class of compounds within Formula Iconsists of those compounds wherein each of R¹, R², R³, R⁴ and R⁵ ishydrido; wherein each of Z¹ and Z² is OH, and wherein the A ring can beeither saturated, partially unsaturated or fully unsaturated (aromatic).

A most highly preferred class of compounds within Formula I consists ofthose compounds wherein each of R¹, R², R³, R⁴ and R⁵ is hydrido;wherein each of Z¹ and Z² is hydroxyl and wherein the A ring is fullyunsaturated (aromatic).

An example of a specific, most highly preferred compound within FormulaI is 5-phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline. This compoundexists as a racemic mixture, as the dextro-isomer and as thelevo-isomer. Also, this compound may be in the form of a salt, includingalkali metal salts such as the sodium salt.

The term "hydrido" denotes a single hydrogen atom (H) which may beattached, for example, to a carbon atom or to an oxygen atom to form anhydroxyl group. Where the term "alkyl" is used, either alone or withinother terms such as "haloalkyl", "aralkyl" and "hydroxyalkyl", the term"alkyl" embraces linear or branched radicals having one to about tencarbon atoms. Preferred alkyl radicals are "lower alkyl" radicals havingone to about five carbon atoms. The term "haloalkyl" embraces radicalswherein any one or more of the carbon atoms is substituted with one ormore halo groups, preferably selected from bromo, chloro and fluoro.Specifically embraced by the term "haloalkyl" are monohaloalkyl,dihaloalkyl and polyhaloalkyl groups. A monohaloalkyl group, forexample, may have either a bromo, a chloro, or a fluoro atom within thegroup. Dihaloalkyl and polyhaloalkyl groups may be substituted with twoor more of the same halo groups, or may have a combination of differenthalo groups. A dihaloalkyl group, for example, may have two bromo atoms,such as a dibromomethyl group, or two chloro atoms, such as adichloromethyl group, or one bromo atom and one chloro atom, such asbromochloromethyl group. Examples of a polyhaloalkyl aretrifluoromethyl, 2,2,2-trifluoroethyl, perfluoroethyl and2,2,3,3-tetrafluoropropyl groups. The term "alkylthio", as representedby the fragment --SR⁵, embraces radicals having a linear or branchedalkyl portion of one to about ten carbon atoms attached to a divalentsulfur atom, such as a methylthio group. The term "alkoxy", asrepresented by the fragment --OR⁵, embraces radicals having a linear orbranched alkyl portion of one to about ten carbon atoms attached to anoxygen atom, such as a methoxy group. The term "aryl" embraces aromaticradicals such as phenyl and naphthyl. The term "aralkyl" embracesaryl-substituted alkyl radicals such as benzyl, diphenylmethyl andtriphenylmethyl. The terms "benzyl" and "phenylmethyl" areinterchangeable.

The term "pharmaceuticaly acceptable salts" embraces forms of a salt ofaddition with a pharmaceutically utilizable acid, either an inorganicacid such as hydrochloric, hydrobromic, hydroiodic, nitric, carbonic,sulfuric or phosphoric acid, or an appropriate organic acid such as analiphatic, cycloaliphatic, aromatic, araliphatic, heterocyclic,carboxylic or alkylsulfonic acid, specific examples of which are formic,acetic, propionic, succinic, glycolic, gluconic, lactic, malic,tartaric, citric, ascorbic, glucuronic, maleic, fumaric, pyruvic,aspartic, glutamic, benzoic, anthranilic, p-hydroxybenzoic, salicylic,phenylacetic, mandelic, embonic (pamoic), methanesulfonic,ethanesulfonic, 2-hydroxyethanesulfonic, panthotenic, benzenesulfonic,toluenesulfonic, sulfanilic, mesylic, cyclohexylaminosulfonic, stearic,alginic, β-hydroxybutyric, malonic, galactaric and galacturonic acid.Also embraced are metallic salts made from aluminium, calcium, lithium,magnesium, potassium, sodium and zinc, and organic salts made frombenzathine (N,N'-dibenzylethylenediamine), chloroprocaine, choline,diethanolamine, ethylenediamine, meglumine (N-methylglucamine) andprocaine.

The compounds of Formula I can possess one or more asymmetric carbonatoms and are thus capable of existing in the form of different, pureoptical isomers as well as in the form of racemic or non-racemicmixtures thereof. All these forms fall within the scope of the presentinvention. The optical isomers can be obtained by resolution of theracemic mixtures according to conventional processes, for example, byformation of diastereomeric salts by treatment with an optically activeacid, such as tartaric, diacetyltartaric, dibenzoyltartaric,ditoluoyltartaric and camphorsulfonic acid, followed by separation ofthe mixture of diastereomers by crystallization and then followed byliberation of the optically active bases from these salts. Separation ofoptical isomers may also be achieved by passing the isomer mixturethrough a chiral chromatography column optimally chosen to maximize theseparation of the enantiomers of the products of the invention orderivatives thereof. Still another available method involves synthesisof covalent stereoisomeric molecules by reacting the compounds of theinvention with an optically pure acid in an activated form or anoptically pure isocyanate. The synthesized diastereoisomers can then beseparated by conventional means such as chromatography, distillation,crystallization or sublimation and submitted to an hydrolytic step whichwill deliver the enantiomerically pure compound. The optically activecompounds according to Formula I can likewise be obtained by utilizingoptically active starting materials. All of these stereoisomers, opticalisomers, diastereomers, as well as mixtures thereof, such as racemicmixtures, are within the scope of the invention.

A therapeutically-active compound of Formula I may be administeredalone, or in a solvent, but is more likely to be included in apharmaceutically-acceptable composition. Such pharmaceuticalcompositions may contain, as active ingredient, at least one compound ofFormula I or its salt of addition with a pharmaceutically utilizableacid, and one or more suitable excipients. These compositions areprepared in such a manner that they can be administered by oral, rectal,parental or local route. The compositions can be solids, liquids or gelforms and may be utilized, according to the administration route, in theform of powders, tablets, lozenges, coated tablets, capsules,granulates, syrups, suspensions, emulsion solutions, suppositories orgels. These compositions can likewise comprise another therapeutic agenthaving an activity similar to or different from that of the compounds ofthe invention.

Other examples of specific compounds of Formula I are listed in Table I:

                  TABLE I                                                         ______________________________________                                        5-Phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline                          hydrochloride;                                                                5-Phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline;                         6-Methyl-5-phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline;                7-Methyl-5-phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline;                8-Methyl-5-phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline;                6-Chloro-5-phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline;                7-Chloro-5-phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline;                8-Chloro-5-phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline;                (D)-5-Phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline;                     (L)-5-Phosphono-3-carboxy-1,2,3,4-tetrahydroisoquinoline;                     5-Phosphono-3-(ethoxycarbonyl)-1,2,3,4-tetrahydroisoquinoline;                5-(Ethyl phosphono)-3-carboxy-1,2,3,4-tetrahydroisoquinoline;                 3-cis-carboxy-5-cis-phosphono-cis-2-azadecalin;                               3-cis-carboxy-5-trans-phosphono-cis-2-azadecalin;                             3-trans-carboxy-5-trans-phosphono-cis-2-azadecalin;                           3-trans-carboxy-5-cis-phosphono-cis-2-azadecalin;                             3-cis-carboxy-5-cis-phosphono-trans-2-azadecalin;                             3-cis-carboxy-5-trans-phosphono-trans-2-azadecalin;                           3-trans-carboxy-5-trans-phosphono-trans-2-azadecalin; and                     3-trans-carboxy-5-cis-phosphono-trans-2-azadecalin.                           ______________________________________                                    

Compounds of Formula I may be prepared in accordance with the followinggeneral procedure: ##STR6##

One process which can be used to synthesize the products of theinvention starts with an ortho toluene derivative of Compound 1 whereeach of R¹, R² and R³ has the values defined previously and L is a goodleaving group such as, for example, halogen, mesylate, tosylate,brosylate and acetate. These ortho toluene derivatives may be treatedwith dialkylphosphites in the presence of a palladium catalyst ortreated first with magnesium in an aprotic anhydrous solvent to form theGrignard reagent which is then reacted further with a chlorodialkylphosphate reagent. The reaction is best achieved by mixingappropriate quantities of the reagents either neat or in a solvant liketoluene, tetrahydrofuran, ether or in a protic solvent in the case ofthe palladium catalyzed reaction, according to the solubility of thereagents, and the reaction temperature can vary from about 0° C. toreflux of the reaction mixture. In the second step of the process, themethyl group of Compound 2 is oxidized to Compound 3. This step is bestachived by treating Compound 2 with an agent able to deliver halogenatoms such as N-bromosuccinimide or N-chlorosuccinimide. The reaction isbest conducted in an halogenated solvent such as chloroform,dichloromethane, tetrachloromethane or trichloroethylene at atemperature between 0° C. and reflux temperature of the solvent, with orwithout irradiation, and in the presence or not of a radical initiatorsuch as azo-bisisobutyronitrile (AIBN). The leaving group is substitutedin the third step with a glycine synthon such as diethylmalonate,acetamidomalonate (X=COOR⁵, Y=CH₃ CO), formamidomalonate (X=COOR⁵,Y=HCO), trifluoroacetamidomalonate (X=COOR⁵, Y=CF₃ CO),methylsulfonamidomalonate (X= COOR⁵, Y=CH₃ SO₂),N-(diphenylmethylene)glycine ethyl ester (X=H, Y=Φ₂ C=) or ethylisocyanoacetate (X=H, Y=:C=). Compound 4 obtained from this reaction mayrequire some transformation of the nitrogen substituent Y. For instance,the formamido, acetamido, isocyano and diphenylmethylene residues can behydrolyzed to the free amine which will be either acylated or sulfonatedto provide a compound more suitable for the experimental conditions ofthe next step. If dietbylmalonate has been used as the glycine synthon,it may be necessary to conduct a mono hydrolysis of the diester bystirring the compound in the presence of one equivalent of an alkalihydroxide such as lithium, sodium or potassium hydroxide at roomtemperature. The acid is carefully transformed into the azido acideither by the mixed anhydride method or by the use of a specific reagentsuch as diphenylphosphoryl azide. The azido acid is transformed into theamine by thermolysis in an aprotic solvent such as toluene and quenchingof the isocyanate formed with dilute HCl.

The cyclization is best conducted by stirring Compound 4 in the presenceof paraformaldehyde or trioxane and methanesulfonic acid in1,2-dichloroethane or in an other chlorinated solvent. When R⁴ is equalto acyl or alkoxycarbonyl, complete hydrolysis of Z¹, Z² and R⁴ can beacheived by an aqueous acid solution, such as 6N HCl or other mineralacid solution. Selective hydrolysis of R⁴ can be achieved in an acidicalcoholic solution. Selective cleavage of Z¹ and Z² can be achieved bycatalytic hydrogenation when Z¹ or Z² is benzyloxy. Various selectivedeprotection schemes are possible depending on the nature of R⁴, Z¹ andZ². When R¹, R² or R³ is hydrolyzable the preferred method ofdeprotection is by catalytic hydrogenation of hydrogenolytically labilegroups.

The perhydroisoquinolines can be prepared by the catalytic hydrogenationof the fully or partially deprotected tetrahydroisoquinolines 5 usingvarious metal catalysts such as Pd, Pt, Ni, Ru and Rh. Partiallyhydrogenated material can be prepared by selective reduction methods,such as the Birch reduction, to obtain dienes followed by selectivecatalytic hydrogenation to obtain the mono-unsaturated products or thefully saturated compound. The advantage in using different methods ofreduction is that different isomers could be obtained among thedifferent racemic mixtures theoretically available.

The following Examples are detailed descriptions of the methods ofpreparation of compounds of Formula I. These detailed preparations fallwithin the scope of, and serve to exemplify, the above described generalprocedures which form part of the invention. These Examples arepresented for illustrative purposes only and are not intended as arestriction on the scope of the invention. All parts are by weightunless otherwise indicated. Most of the commercially-available startingmaterials were obtained from Aldrich Chemical Co., Milwaukee, Wis.

EXAMPLE I ##STR7##

Diethyl 2-methylphenylphosphonate (6.84 gm) and N-bromosuccinimide (NBS)(5.87 gm) were combined in CCl₄ (60 mL). A small amount ofazo-bisisobutyronitrile was added and the mixture was heated to reflux.After 6 hours, the NBS had been completely consumed and the orangecolored reaction mixture had become a pale yellow. The reaction mixturewas cooled to room temperature and the insoluble succinimide removed byfiltration. Removal of the solvent on a rotary evaporator afforded ayellow oil. The oil was chromatographed on silica gel (125 gm) elutingwith ethyl acetate. The appropriate fractions were pooled andconcentrated to afford the product as a colorless oil. ##STR8## Sodium(82 mg) was dissolved in anhydrous ethanol (6 ml) under a nitrogenatmosphere. Diethyl formamidomalonate (725 mg) was then added withstirring. The reaction mixture became homogeneous and then a precipitatebegan to form. The reaction mixture was briefly heated to reflux andthen allowed to cool. The 2-(diethylphosphono)benzyl bromide (1 gm) wasthen added and the reaction allowed to stir at room temperature for 20hours. The reaction mixture was partitioned between H₂ O (30 ml) and Et₂O (30 ml). The lower aqueous layer was extracted with fresh Et₂ O (30ml) and the combined Et₂ O layers washed once with saturated NaCl (30ml). The Et₂ O layer was then dried (MgSO₄) and concentrated to an oil.The oil was chromatographed on silica gel using ethyl acetate as theeluting solvent. The appropriate fractions were pooled and concentratedto afford the product as a clear oil. ##STR9## Diethyl2-(2-(diethylphosphono)benzyl)formamidomalonate (430 mg) was combinedwith paraformaldehyde (31 mg) and acetic anhydride (94 μl) in1,2-dichloroethane (3.6 ml) containing methanesulfonic acid (0.4 ml) andallowed to stir at room temperature for 7 days. The reaction mixture wasdiluted with Et₂ O (25 ml) and extracted with H₂ O (10 ml). The H₂ Olayer was extracted with Et₂ O (25 ml) and the combined Et₂ O layersdried (MgSO₄) and concentrated to an oil. The oil was chromatographed onsilica gel (50 gm) equilibrated with CH₂ Cl₂. The column was eluted withCH₂ Cl₂ (100 ml), 1% EtOH/CH₂ Cl₂ (200 ml), and then the eluent was heldat 2% EtOH/CH₂ Cl₂. Fractions of about 10 ml were collected. A few minorimpurities eluted followed by the product in fractions 71-79 andunreacted starting material in fractions 82- 92. The appropriatefractions were pooled and concentrated to an oil.

EXAMPLE II ##STR10##

N-Formyl-3-bis(ethoxycarbonyl)-5-(diethylphosphono)-1,2,3,4-tetrahydroisoquinoline(100 mg) was combined with 6N HCl (20 ml) and heated to reflux for 18hours. The solvent was removed on a rotary evaporator and the resultingwhite solid dissolved in H₂ O (20 ml) and reconcentrated. This processwas repeated with ethanol and the final product dried in vacuo.

Elemental Analysis

    ______________________________________                                                   Theory + H.sub.2 O                                                                      Found                                                    ______________________________________                                        C            38.54       38.10                                                H            4.85        4.90                                                 N            4.49        4.34                                                 ______________________________________                                    

¹ H NMR (D₂ O) δ* 3.38 (m,1H), 3.78 (m,1H), 4.40 (t,1H), 4.44 (m,2H),7.36 (m,2H), 7.75 (m,1H).

BIOLOGICAL EVALUATION Binding Assays

[Pullan, L. M., Olney, J. W., Price, M. T., Compton, R. P., Hood, W. F.,Michel J., Monahan J. B., "Excitatory Amino Acid Receptor Potency andSubclass Specificity of Sulfur-Containing Amino Acids", Journal ofNeurochemistry, 49 1301-1307, (1987)].

Synaptic plasma membranes (SPM) were prepared as previously-described[Monahan, J. B. and Michel, J., "Identification and Characterization ofan N-methyl-D-aspartate-specific L[³ H]glutamate Recognition Site inSynaptic Plasma Membranes, J. Neurochem., 48, 1699-1708 (1987)]. The SPMwere stored at a concentration of 10-15 mg/ml in 0.32M sucrose, 0.5 mMEDTA, 1 mM MgSO₄, 5 mM Tris/SO₄, pH 7.4, under liquid nitrogen. Theidentity and purity of the subcellular fractions were confirmed by bothelectron microscopy and marker enzymes. Protein concentrations weredetermined by using a modification of the method of Lowry [Ohnishi, S.T. and Barr, J. K., "A Simplified Method of Quantitating Proteins usingthe Biuret and Phenol Reagents", Anal. Biochem., 86, 193-197 (1978)].

The SPM were treated identically for the [³ H]AMPA (QUIS), [³ H]kainateand sodium-dependent L-[³ H]glumatate binding assays. The SPM werethawed at room temperature, diluted twenty-fold with 50 mM Tris/acetate,pH 7.4, incubated at 37° C. for 30 minutes, and centrifuged at 100,000 gfor 15 minutes. The dilution, incubation and centrifugation wererepeated a total of three times.

Prior to use in the NMDA-specific L-[³ H]glutamate binding assay, theSPM were thawed, diluted twenty fold with 50 mM Tris/acetate (pH 7.4containing 0.04% (v/v) Triton X-100), incubated for 30 minutes at 37° C.and centrifuged as described above. The Triton X-100 treated membraneswere washed with 50 mM Tris/acetate (pH 7.4) and centrifuged at 100,000g for 15 minutes a total of four times.

The basic procedure for the receptor subclass binding assays wassimilar. This general method involved adding the radioligand (12.5 nML-[³ H] glutamate; 0.5 nM [³ H]kainate or 10 nM [³ H]AMPA) to theappropriate concentration of the test compound and initiating the assayby the addition of ice cold synaptic plasma membranes (0.2-0.45 mg). Thebinding assays were performed in 1.5 mL centrifuge tubes with the totalvolume adjusted to 1.0 mL. Additions of test compounds were made in 50mM Tris/acetate (pH 7.4) and incubations were carried out at 0°-4° C.The incubation time for each of the NMDA and the AMPA binding assays was10 minutes, for the kainate binding assay 60 minutes and for thesodium-dependent glutamate binding assay 15 minutes. The AMPA bindingassay contained 100 mM KSCN and the sodium-dependent glutamate bindingassay contained 150 mM sodium acetate in addition to the previouslydescribed reagents.

To terminate the incubation, the samples were centrifuged for 15 minutesat 12,000 g and 4° C. in a Beckman Microfuge 12. The supernatant wasaspirated and the pelleted membranes dissolved in Beckman BTS-450 tissuesolubilizer for a minimum of 6 hours at room temperature. Beckman MPscintillation cocktail containing 7 mL/1 acetic acid was then added andthe samples counted on a Beckman LS 5800 or 3801 liquid scintillationcounter with automatic corrections for quenching and countingefficiency.

Nonspecific binding was defined as the residual binding in the presenceof either excess L-glutamate (0.1-0.4 mM), kainate (0.01 mM), or NMDA(0.5 mM), and was 15-25% of the total binding in the NMDA binding assay,19-27% in the AMPA binding assay, 20-30% in the kainate binding assayand 10-15% in the sodium-dependent binding assay. Radioligand binding tothe synaptic plasma membranes was analyzed using Scatchard and Hilltransformations and the K_(i) values of the compounds determined usinglogit-log transformations. Calculations and regression analysis wereperformed using templates developed for Lotus 1, 2, 3 as previouslydescribed [Pullan, L. M. "Automated Radioligand Receptor BindingAnalysis with Templates for Lotus", Computer Appln. Biosci., 3 131(1987)]. Binding results are reported in Table II for example compoundsof the invention. Included in Table II are binding data forD,L-AP7[D,L-2-amino-7-phosphonoheptanoic acid].

                  TABLE II                                                        ______________________________________                                        RECEPTOR BINDING DATA                                                                 Binding (μM)                                                       Compound  NMDA          KA      Quis                                          ______________________________________                                        D,L-AP7   5.4           >300    >300                                          D-AP7     4.0           >300    >300                                          Ex. II    1.6           ˜300                                                                            >300                                          ______________________________________                                    

TCP Modulation Assay

The effect on the TCP (1-[1-(2-thienyl)-cyclohexyl]piperidine) bindingwas measured in rat brain synaptic membranes (SPM) prepared aspreviously described [J. B. Monahan & J. Michel; J. Neurochem.48:1699-1708 (1987)]. Prior to their use in the binding assay, frozenSPM were thawed, diluted twenty fold with 50 mM Tris/acetate (pH 7.4containing 0.04% (v/v) Triton X-100), incubated for 30 min. at 37° C.and centrifuged at 95,000 xg for 15 min. The Triton X-100 treated SPMwere washed with 5 mM Tris/HCl, pH 7.4 and centrifuged a total of sixtimes. The compound of Example II was incubated at differentconcentrations with SPM (0.2-0.4 mg protein) and 2 nM tritiated TCP, ina total volume of 0.5 ml of 5 mM Tris/HCl buffer pH 7.4 at 25° C. for 60min. The samples were filtered through glass fiber filters (Schleicher &Schuell #32) which have been pretreated with 0.05% (v/v)polyethylenimine, washed 4 times with 2 ml of ice-cold 5 mM Tris/HClbuffer, and then counted on a Beckman LS 5800 liquid scintillationcounter with automatic corrections for quenching and countingefficiency. Inhibition of TCP binding was measured as a decrease in thebinding in the presence of 0.05 mM L-glutamate. Non-specific binding wasdefined as the residual binding in the presence of 60 mM phencyclidine.

Result

The compound of Example II inhibits 64% of TCP binding at 5 μM and 91%at 50 μM.

Administration of compounds within Formula I to humans can be by anytechnique capable of introducing the compounds into the bloodstream of ahuman patient, including oral administration, and by intravenous,intramuscular and subcutaneous injections.

Compounds indicated for prophylactic therapy will preferably beadministered in a daily dose generally in a range from about 0.1 mg toabout 100 mg per kilogram of body weight per day. A more preferreddosage will be a range from about 1 mg to about 100 mg per kilogram ofbody weight. Most preferred is a dosage in a range from about 1 to about50 mg per kilogram of body weight per day. A suitable dose can beadministered, in multiple sub-doses per day. These sub-doses may beadministered in unit dosage forms. Typically, a dose or sub-dose maycontain from about 1 mg to about 100 mg of active compound per unitdosage form. A more preferred dosage will contain from about 2 mg toabout 50 mg of active compound per unit dosage form. Most preferred is adosage form containing from about 3 mg to about 25 mg of active compoundper unit dose.

The active compound is usually administered in apharmaceutically-acceptable formulation, although in some acute-caresituations a compound of Formula I may be administered alone. Suchformulations may comprise the active compound together with one or morepharmaceutically-acceptable carriers or diluents. Other therapeuticagents may also be present in the formulation. Apharmaceutically-acceptable carrier or diluent provides an appropriatevehicle for delivery of the active compound without introducingundesirable side effects. Delivery of the active compound in suchformulations may be by various routes including oral, nasal, topical,buccal and sublingual, or by parenteral administration such assubcutaneous, intramuscular, intravenous and intradermal routes.

Formulations for oral administration may be in the form of capsulescontaining the active compound dispersed in a binder such as gelatin orhydroxypropylmethyl cellulose, together with one or more of a lubricant,preservative, surface-active or dispersing agent. Such capsules ortablets may contain controlledrelease formulation as may be provided ina dispersion of active compound in hydroxypropylmethyl cellulose.

Formulations for parenteral administration may be in the form of aqueousor non-aqueous isotonic sterile injection solutions or suspensions.These solutions and suspensions may be prepared from sterile powders orgranules having one or more of the carriers or diluents mentioned foruse in the formulations for oral administration.

Although this invention has been described with respect to specificembodiments, the details of these embodiments are not to be construed aslimitations. Various equivalents, changes and modifications may be madewithout departing from the spirit and scope of this invention, and it isunderstood that such equivalent embodiments are part of this invention.

What is claimed is:
 1. A compound of the formula ##STR11## wherein eachof R¹ through R³ is independently selected from hydrido, alkyl,haloalkyl, halo, cyano, nitro and groups represented by --OR⁵, SR⁵,##STR12## wherein R⁵ is selected from hydrido, alkyl, aryl and aralkyl;and wherein R⁴ is hydrido or formyl;and wherein each of z¹ and Z² isindependently selected from --OR⁵, SR⁵, ##STR13## wherein R⁵ is definedas before; and wherein the A ring is saturated; or apharmaceutically-acceptable salt thereof.
 2. A compound of the formula##STR14## wherein each of R¹, R² and R³ is independently selected fromhydrido, alkyl containing from one to ten carbon atoms, haloalkylwherein the alkyl portion thereof contains from one to ten carbon atoms,halo, cyano, nitro and groups represented by --OR⁵ and --SR⁵ ;wherein R⁵is selected from hydrido, alkyl containing from one to ten carbon atoms,aryl containing from six no ten carbon atoms and aralkyl wherein thealkyl portion thereof contains from one to ten carbon atoms and the arylportion thereof contains from six to ten carbon atoms; and wherein eachof Z¹ and Z² is independently selected from ##STR15## wherein R⁴ ishydrido or formyl; wherein R⁵ is defined as before; and wherein the Aring is saturated; or a pharmaceutically-acceptable salt thereof. 3.Compound of claim 2 wherein each of R¹, R² and R³ is hydrido. 4.Compound of claim 3 wherein each of R¹ to R⁵ is hydrido; and whereineach of Z¹ and Z² is hydroxyl.
 5. Compound of claim 2 selected from thegroup consisting of:3-cis-carboxy-5-cis-phosphono-cis-2-azadecalin;3-cis-carboxy-5-trans-phosphono-cis-2-azadecalin;3-trans-carboxy-5-trans-phosphono-cis-2-azadecalin;3-trans-carboxy-5-cis-phosphono-cis-2-azadecalin;3-cis-carboxy-5-cis-phosphono-trans-2-azadecalin;3-cis-carboxy-5-trans-phosphono-trans-2-azadecalin;3-trans-carboxy-5-trans-phosphono-trans-2-azadecalin; and3-trans-carboxy-5-cis-phosphono-trans-2-azadecalin.
 6. A pharmaceuticalcomposition comprising a therapeutically-effective amount of a compoundand a pharmaceutically-acceptable carrier or diluent, said compoundselected from a family of compounds of the formula ##STR16## whereineach of R¹, R² and R³ is independently selected from hydrido, alkylcontaining from one to ten carbon atoms, haloalkyl wherein the alkylportion thereof contains from one to ten carbon atoms, halo, cyano,nitro and groups represented by --OR⁵ and --SR⁵ ; wherein R⁵ is selectedfrom hydrido, alkyl containing from one to ten carbon atoms, arylcontaining from six no ten carbon atoms and aralkyl wherein the alkylportion thereof contains from one to ten carbon atoms and the arylportion contains from six to ten carbon atoms;and wherein each of Z¹ andZ² is independently selected from --OR⁵, SR⁵, ##STR17## wherein R⁴ ishydrido or formyl; wherein R⁵ is defined as before; and wherein the Aring is saturated; or a pharmaceutically-acceptable salt thereof.
 7. Thecomposition of claim 6 wherein each of R¹, R² and R³ is hydrido.
 8. Thecomposition of claim 7 wherein each of R¹ to R⁵ is hydrido; and whereineach of Z¹ and Z² is hydroxyl.
 9. The composition of claim 6 whereinsaid compound is selected from the group consistingof;3-cis-carboxy-5-cis-phosphono-cis-2-azadecalin;3-cis-carboxy-5-trans-phosphono-cis-2-azadecalin;3-trans-carboxy-5-trans-phosphono-cis-2-azadecalin;3-trans-carboxy-5-cis-phosphono-cis-2-azadecalin;3-cis-carboxy-5-cis-phosphono-trans-2-azadecalin;3-cis-carboxy-5-trans-phosphono-trans-2-azadecalin;3-trans-carboxy-5-trans-phosphono-trans-2-azadecalin; and3-trans-carboxy-5-cis-phosphono-trans-2-azadecalin.
 10. A method tocontrol excitatory amino acid induced neurotoxic injury mediated by anNMDA receptor in meals, which method comprises treating a mammalsusceptible to neurologic injury with an effective amount of a compoundof the formula ##STR18## wherein each of R¹, R² and R³ is independentlyselected from hydrido, alkyl containing from one to ten carbon atoms,haloalkyl wherein the alkyl portion thereof contains from one to tencarbon atoms, halo, cyano, nitro and groups represented by --OR⁵ and--SR⁵ ;wherein R⁵ is selected from hydride, alkyl containing from one toten carbon atoms, aryl containing from six to ten carbon atoms andaralkyl wherein the alkyl portion thereof contains from one to tencarbon atoms and the aryl portion contains from six to ten carbon atoms;and wherein each of Z¹ and Z² is independently selected from --OR⁵, SR⁵,##STR19## wherein R⁴ is hydrido or formyl; wherein R⁵ is defined asbefore; and wherein the A ring is saturated; or apharmaceutically-acceptable salt thereof.
 11. The method of claim 10wherein each of R¹, R² and R³ is hydrido.
 12. The method of claim 11wherein each of R¹ to R⁵ is hydrido; wherein each of Z¹ and Z² ishydroxyl.
 13. The method of claim 10 wherein said compound is selectedfrom the group consistingof:3-cis-carboxy-5-cis-phosphono-cis-2-azadecalin;3-cis-carboxy-5-trans-phosphono-cis-2-azadecalin;3-trans-carboxy-5-trans-phosphono-cis-2-azadecalin;3-trans-carboxy-5-cis-phosphono-cis-2-azadecalin;3-cis-carboxy-5-cis-phosphono-trans-2-azadecalin;3-cis-carboxy-5-trans-phosphono-trans-2-azadecalin;3-trans-carboxy-5-trans-phosphono-trans-2-azadecalin; and3-trans-carboxy-5-cis-phosphono-trans-2-azadecalin.
 14. Compound ofclaim 5 which is 3-cis-carboxy-5-cis-phosphono-cis-2-azadecalin or apharmaceutically-acceptable salt thereof.
 15. The composition of claim 9wherein said compound is 3-cis-carboxy-5-cis-phosphono-cis-2-azadecalinor a pharmaceutically-acceptable salt thereof.
 16. The method of claim13 wherein said compound is3-cis-carboxy-5-cis-phosphono-cis-2-azadecalin or apharmaceutically-acceptable salt thereof.